PROJECT SUMMARY/ABSTRACT Despite advancements in combination chemotherapy, targeted therapies, and immunotherapies, most advanced cancers remain incurable. Deciphering the mechanisms that govern the initiation and progression from early to advanced cancer will facilitate the development of novel strategies for prevention and therapy. For cancers to arise, cells must acquire the capacity for sustained proliferation while overcoming both intrinsic evolutionary constraints and constraints imposed by the host environment. These properties may be attained through the sequential acquisition of genetic mutations in proto-oncogenes and tumor suppressor genes. Using genetically-engineered mouse models that closely recapitulate human cancers, we have shown that mutations are permissive, but insufficient, to drive clonal cancer evolution, consistent with the need for additional cell-autonomous adaptations. Furthermore, we have demonstrated that host adaptations in endocrine hormone production in response to environmental stresses, such as obesity, may facilitate tumor progression in the absence of new mutations. We propose to define the molecular basis for the tumor cell intrinsic and extrinsic adaptations observed in these models. By tracing genetically-distinct subclonal populations within tumors at single cell resolution and applying new technologies and analytic tools for single cell analysis, we will discover and functionally validate transcriptional and epigenetic adaptations within tumor cells that cooperate with mutations to promote clonal cancer evolution. In complementary studies, we will use both molecular and proteomic approaches to fully characterize hormonal adaptations to obesity, a major risk factor for many cancers, and how alterations in endocrine hormone production may influence cancer progression using in vitro and in vivo preclinical models. Finally, we will determine whether the adaptations observed in these models occurs in patients by evaluating the expression of the discovered genes and hormones in human biospecimens. Together, our proposed studies not only have the potential to fill an important gap in knowledge by defining critical adaptations for cancer development but also discover new targets and approaches for furthering cancer therapy.